It is well known that incretins released by enteroendocrine cells increase insulin secretion from the pancreatic beta-cell in response to oral nutrients. Glucagon-Like Peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are the two major incretins most extensively studied to date. In addition to regulating insulin release, incretins increase -cell mass and lower plasma glucose via extra-pancreatic effects. Thus, they are important for maintaining normal glucose tolerance. Recently approved therapies in widespread use for treating type 2 diabetes mellitus (T2DM) modulate GLP-1 levels or action. However, humans with T2DM are resistant to the action of GIP and this peptide has not been used to treat T2DM. Our preliminary studies in transgenic mice engineered to lack K cells have shown xenin-25, a second peptide product of the K cell, enhances insulin secretory responses to GIP. The mechanism responsible is novel because xenin-25 does not act directly on the pancreatic beta-cell, but stimulates acetylcholine release from cholinergic neurons that innervate the islets thereby activating muscarinic receptors on the pancreatic -cell. This unexpected finding is of interest in understanding the physiology of insulin secretion, and suggests a novel approach to enhancing endogenous insulin secretion in T2DM. We now propose to build on these preliminary data and determine if xenin-25 plays a previously unappreciated role in regulating insulin secretion, beta-cell mass, and as a potential therapeutic agent for T2DM. Only 1 published study has involved the administration of xenin-25 to humans and effects on insulin secretion and glucose tolerance were not described. If xenin-25 does have effects on insulin secretion, this would not only define an unanticipated element of insulin secretory biology but would suggest a new treatment for T2DM. Our studies are thus completely novel. Our long-term goal is to understand how peptides produced by K cells regulate -cell function. Our central hypothesis is that xenin-25 can amplify the effects of GIP on the beta-cell to increase the insulin secretory response to glucose and also to improve beta-cell mass. This project has 2 aims. First, we will define the role of xenin-25 in regulating insulin secretion in humans with normal glucose tolerance, impaired glucose tolerance, and T2DM by measuring insulin secretory rates during hyperglycemic clamps and graded glucose infusions with GIP versus GIP plus xenin-25. Because GIP production/release is normal in T2DM, we will also define plasma xenin-25 levels in humans and determine if they are perturbed in T2DM. Second, we will define the effects and mechanism of action of xenin-25 on beta-cell function in mice by studying the roles of the vagus nerve and M3 muscarinic receptors on pancreatic 2-cells in mediating the effects of xenin-25 on GIP-mediated insulin release. We will also determine if xenin-25 potentiates the effects of GIP to enhance insulin production and beta-cell mass.